1. Field of the Invention
The present invention relates generally to microscopes, and in particular, to a method, apparatus, and article of manufacture for a differential interference contrast (DIC) microscope and/or light field profiler based on Young's interference. In addition, the present invention provides a method, apparatus, and article of manufacture for an optofluidic microscope that is surface plasmon assisted. Both methods are related in that they make novel usage of interference between holes and/or structured holes to enable higher sensitivity and a different contrast method in on-chip microscopy imaging and light field profiling.
2. Description of the Related Art
(Note: This application references a number of different publications as indicated throughout the specification by reference numbers enclosed in brackets, e.g., [x]. A list of these different publications ordered according to these reference numbers can be found below in the section entitled “References.” Each of these publications is hereby incorporated by reference in its entirety for all purposes.)
Differential Interference Contrast (DIC) Microscopes
A major problem of imaging transparent specimens with conventional microscopes is that it can be difficult to elicit contrast, because the imaging technique is solely based on the amplitude information provided by the sample. This difficulty is especially true for most of the biological samples. Therefore phase information, if measured, will improve the imaging contrast dramatically. Differential interference contrast (DIC) microscope performs admirably in this respect by rendering excellent phase contrast in transparent specimens, and is widely used in biology and clinical laboratories.
DIC microscopes are a beam-shearing interference system [DIC1]. A reference beam is sheared by a very small distance with respect to a sample beam. The phase difference between the reference beam and the sample beam after they pass two adjacent spots of the specimen provides the differential phase contrast of the specimen. Since DIC microscopy is an interference-based technique, it can distinguish minuscule amount of phase differences and identify small changes in the sample's refractive index.
Prior art DIC microscopes have some disadvantages. Firstly, prior art DIC microscopes are very expensive instruments, as many complicated and expensive optical components are required to manipulate the light. Secondly, the lateral resolution of current DIC microscopes is determined by the spot size of the objective lens of the DIC microscope, which has a diffraction limit. The small sheared distance between the reference beam and the sample beam is usually tuned to be slightly smaller than this spot size.
Microfluidics
Recent developments in microfluidics have brought forth a variety of new devices that can potentially revolutionize traditional biomedical and chemical experiments [MICRO2-MICRO7]. One such new device is the optofluidic microscope (OFM) described in U.S. patent application Ser. No. 11/686,095, filed on Mar. 14, 2007, by Changhuei Yang and Demetri Psaltis, entitled “OPTOFLUIDIC MICROSCOPE DEVICE,” which is incorporated by reference herein. The OFM fuses the advantage of optical imaging in providing high resolution and the advantages of microfluidics, such as low cost and high throughput. Further, OFM's application in nematode imaging and phenotyping has been reported [MICRO8].
FIGS. 1A and 1B illustrate an OFM device that consists of an opaque metallic film with an etched array of submicron holes. The metallic film is bonded to the floor of a PDMS (polydimethylsiloxane) microfluidic chip. The hole array is oriented at a small angle relative to the micro-channel (FIG. 1B). As a biological sample passes over the nanohole array (as indicated by the flow direction), each individual hole will take a line scan of the target. The sample is illuminated based on the illuminated direction indicated. FIG. 1B illustrates the top view of the OFM wherein α denotes the isolated hole and β denotes the corresponding hole that scans the same line on the target as hole α does.
The OFM was used to image and perform nematode phenotype characterization of Wild type C. elegans and dpy-24 mutants at their first larval stages as illustrated in FIG. 2. FIG. 2(a) illustrates the OFM image of the wild-type C elegans larvae at the first larval stage. FIG. 2(b) illustrates the OFM image of a dpy-24 mutant. FIG. 2(c) illustrates the aspect ratio of wild-type larvae and dpy-24 mutants.
The resolution limit of OFM and any other nanohole based sensors is given by the hole size. Although smaller hole size gives better optical resolution, the optical transmission through the nanohole will be dramatically reduced [MICRO9 and MICRO10]. The weak transmission signal may be buried by electronic noise or background noise, which can make an isolated subwavelength hole less desirable for optical imaging applications. Although powerful lasers can help to increase the total transmission through the nanohole, high-intensity light may also have adverse effects on the biological samples. Therefore, new schemes that can either enhance the optical transmission of a nanohole or improve the sensitivity of the detection are desirable.
In view of the above, what is needed is the ability to overcome the disadvantages of the prior art DIC microscopes and to improve the capability and sensitivity of optofluidic microscopes.